Notes

  • Unclassified reads were excluded
  • Taxa with total reads number of 10 or lower were excluded
  • Eukaryotes were excluded from the analyis

Import Exp data

MCM D6300

MCM D6331

FR D6323

Canis lupus

Homo sapiens

Merge

Filter Exp data

Filter samples based on total read count

Fix sample names

Gold Standard Data

Exclude (truly) unclassified reads

Convert NA to ‘missing’ in the taxonomy table (to avoid errors)

Number of NAs fo each rank level: 

## superkingdom       phylum        class        order       family        genus 
##            0            0            0            0            0            0 
##      species   Gram.Stain 
##            0            7

‘missing’ means that there is no associated taxon level for the given taxon, but the read itself was classified.
This should be considered when glomerating taxa, as ‘missing’ taxa can be derived from different lineages.
To avoid this, the glomerating should be done using: ignore_lineage=F

Filter for taxa below threshold

Taxon IDs

## using species as taxon identifier

Fix genera

## Glomerating all hits from: Veillonella to: Veillonella rogosae
## Glomerating all hits from: Faecalibacterium to: Faecalibacterium prausnitzii
## Glomerating all hits from: Prevotella to: Prevotella corporis
## Glomerating all hits from: Roseburia to: Roseburia hominis

Fix species

## Glomerating all hits from: Bacteroides fragilis to: Bacteroides fragilis
## Glomerating all hits from: Bacteroides fragilis CAG:47 to: Bacteroides fragilis
## Glomerating all hits from: Bacteroides fragilis_A to: Bacteroides fragilis

Fix sample names from ‘-’

Merge exp data with GS

Filter Taxa for Gold Standard

These are the taxa that were found in the experimental data
species taxid superkingdom phylum class order family genus count Gram Stain EXP_sum
Bacillus subtilis 1423 Bacteria Firmicutes Bacilli Bacillales Bacillaceae Bacillus 1740000 + 2822873.2
Enterococcus faecalis 1351 Bacteria Firmicutes Bacilli Lactobacillales Enterococcaceae Enterococcus 990000 + 1451302.3
Escherichia coli 562 Bacteria Proteobacteria Gammaproteobacteria Enterobacterales Enterobacteriaceae Escherichia 1010000 - 1450435.4
Limosilactobacillus fermentum 1613 Bacteria Firmicutes Bacilli Lactobacillales Lactobacillaceae Limosilactobacillus 1840000 + 2345749.4
Listeria monocytogenes 1639 Bacteria Firmicutes Bacilli Bacillales Listeriaceae Listeria 1410000 + 1850634.6
Pseudomonas aeruginosa 287 Bacteria Proteobacteria Gammaproteobacteria Pseudomonadales Pseudomonadaceae Pseudomonas 420000 - 445095.9
Salmonella enterica 28901 Bacteria Proteobacteria Gammaproteobacteria Enterobacterales Enterobacteriaceae Salmonella 1040000 - 1210133.4
Staphylococcus aureus 1280 Bacteria Firmicutes Bacilli Bacillales Staphylococcaceae Staphylococcus 1550000 + 2060483.9

Add Gram Stain Info

Omit Eukaryotes

Relative abundance of taxa at each taxonomic level

Parameters and Settings

Correlation

PERMANOVA model

Gram Stain level

Barplot of abundances

Phylum level

Barplot of abundacnes

MAASLIN2 - Number of Significantly Different Taxa Among FR, H. sapiens and C. lupus

Barplot of Coefficients

Heatmap of Significant Associations (−log(qval) x sign(coeff))

PERMANOVA

Df SumOfSqs R2 F Pr(>F)
primer 2 0.2220239 0.1815434 2.661743 0.069
Residual 24 1.0009559 0.8184566 NA NA
Total 26 1.2229798 1.0000000 NA NA

Class level

Barplot of abundances

MAASLIN2 - Number of Significantly Different Taxa Among FR, H. sapiens and C. lupus

Barplot of Coefficients

Heatmap of Significant Associations (−log(qval) x sign(coeff))

PERMANOVA

Df SumOfSqs R2 F Pr(>F)
primer 2 0.2551431 0.1460105 2.051694 0.047
Residual 24 1.4922872 0.8539895 NA NA
Total 26 1.7474303 1.0000000 NA NA

Order level

Barplot of abundances

MAASLIN2 - Number of Significantly Different Taxa Among FR, H. sapiens and C. lupus

Barplot of Coefficients

Heatmap of Significant Associations (−log(qval) x sign(coeff))

### PERMANOVA

Df SumOfSqs R2 F Pr(>F)
primer 2 0.2509976 0.1403119 1.958551 0.056
Residual 24 1.5378566 0.8596881 NA NA
Total 26 1.7888542 1.0000000 NA NA

Family level

Barplot of abundances

MAASLIN2 - Number of Significantly Different Taxa Among FR, H. sapiens and C. lupus

Barplot of Coefficients

Heatmap of Significant Associations (−log(qval) x sign(coeff))

PERMANOVA

Df SumOfSqs R2 F Pr(>F)
primer 2 0.3283757 0.1074244 1.444239 0.005
Residual 24 2.7284326 0.8925756 NA NA
Total 26 3.0568083 1.0000000 NA NA

Genus level

Barplot of abundances

MAASLIN2 - Number of Significantly Different Taxa Among FR, H. sapiens and C. lupus

Barplot of Coefficients

Heatmap of Significant Associations (−log(qval) x sign(coeff))

PERMANOVA

Df SumOfSqs R2 F Pr(>F)
primer 2 0.2487811 0.0619096 0.7919443 0.014
Residual 24 3.7696752 0.9380904 NA NA
Total 26 4.0184563 1.0000000 NA NA

Species level

Barplot of abundances

MAASLIN2 - Number of Significantly Different Taxa Among FR, H. sapiens and C. lupus

Barplot of Coefficients

Heatmap of Significant Associations (−log(qval) x sign(coeff))

PERMANOVA

Df SumOfSqs R2 F Pr(>F)
primer 2 0.2997329 0.0538113 0.6824602 0.001
Residual 24 5.2703370 0.9461887 NA NA
Total 26 5.5700699 1.0000000 NA NA

Summary of Similarity of FR Taxa to Homo sapiens and Canis lupus

Summary of PERMANOVAs on Different Rank Levels

Summary of Detection and Correlation Statistics for Zymo MCM D6300, D6331 and D6323

Detection Statistics

##                                  sample     N
##                                  <char> <int>
## 1: PRMT_FM1_FR_MONT_20240807_bc25_D6323    60
## 2: PRMT_FM2_FR_MONT_20240807_bc26_D6323    60
## 3: PRMT_FM3_FR_MONT_20240807_bc27_D6323    60
## 4:  PRMT_FO1_FR_ONT_20240807_bc10_D6323    60
## 5:  PRMT_FO2_FR_ONT_20240807_bc11_D6323    60
## 6:  PRMT_FO3_FR_ONT_20240807_bc12_D6323    60
## 7:   PRMT_FP1_FR_PB_20240807_bc40_D6323    60
## 8:   PRMT_FP2_FR_PB_20240807_bc41_D6323    60
## 9:   PRMT_FP3_FR_PB_20240807_bc42_D6323    60
##                                   sample     N
##                                   <char> <int>
## 1: PRMT_MM1_MCS_MONT_20240807_bc19_D6300    60
## 2: PRMT_MM2_MCS_MONT_20240807_bc20_D6300    60
## 3: PRMT_MM3_MCS_MONT_20240807_bc21_D6300    60
## 4:  PRMT_MO1_MCS_ONT_20240807_bc04_D6300    60
## 5:  PRMT_MO2_MCS_ONT_20240807_bc05_D6300    60
## 6:  PRMT_MO3_MCS_ONT_20240807_bc06_D6300    60
## 7:   PRMT_MP1_MCS_PB_20240807_bc34_D6300    60
## 8:   PRMT_MP2_MCS_PB_20240807_bc35_D6300    60
## 9:   PRMT_MP3_MCS_PB_20240807_bc36_D6300    60
##                                   sample     N
##                                   <char> <int>
## 1: PRMT_GM1_GMS_MONT_20240807_bc22_D6331    60
## 2: PRMT_GM2_GMS_MONT_20240807_bc23_D6331    60
## 3: PRMT_GM3_GMS_MONT_20240807_bc24_D6331    60
## 4:  PRMT_GO1_GMS_ONT_20240807_bc07_D6331    60
## 5:  PRMT_GO2_GMS_ONT_20240807_bc08_D6331    60
## 6:  PRMT_GO3_GMS_ONT_20240807_bc09_D6331    60
## 7:   PRMT_GP1_GMS_PB_20240807_bc37_D6331    60
## 8:   PRMT_GP2_GMS_PB_20240807_bc38_D6331    60
## 9:   PRMT_GP3_GMS_PB_20240807_bc39_D6331    60
statistic N
True_positives 162
False_positives 162
False_negatives 162
Precision 162
Recall 162
F1 162
F0.5 162
Chi_stat 162
Chisq.test.p_value 162
Chisq.test.sign 162
GS_version N
D6331 540
D6300 540
D6323 540

Chi^2

F1, Precision and Recall

Correlation of Abundance Values between Theoretical and Experimental

##                                  sample     N
##                                  <char> <int>
## 1: PRMT_FM1_FR_MONT_20240807_bc25_D6323     3
## 2: PRMT_FM2_FR_MONT_20240807_bc26_D6323     3
## 3: PRMT_FM3_FR_MONT_20240807_bc27_D6323     3
## 4:  PRMT_FO1_FR_ONT_20240807_bc10_D6323     3
## 5:  PRMT_FO2_FR_ONT_20240807_bc11_D6323     3
## 6:  PRMT_FO3_FR_ONT_20240807_bc12_D6323     3
## 7:   PRMT_FP1_FR_PB_20240807_bc40_D6323     3
## 8:   PRMT_FP2_FR_PB_20240807_bc41_D6323     3
## 9:   PRMT_FP3_FR_PB_20240807_bc42_D6323     3
##                                   sample     N
##                                   <char> <int>
## 1: PRMT_MM1_MCS_MONT_20240807_bc19_D6300     3
## 2: PRMT_MM2_MCS_MONT_20240807_bc20_D6300     3
## 3: PRMT_MM3_MCS_MONT_20240807_bc21_D6300     3
## 4:  PRMT_MO1_MCS_ONT_20240807_bc04_D6300     3
## 5:  PRMT_MO2_MCS_ONT_20240807_bc05_D6300     3
## 6:  PRMT_MO3_MCS_ONT_20240807_bc06_D6300     3
## 7:   PRMT_MP1_MCS_PB_20240807_bc34_D6300     3
## 8:   PRMT_MP2_MCS_PB_20240807_bc35_D6300     3
## 9:   PRMT_MP3_MCS_PB_20240807_bc36_D6300     3
##                                   sample     N
##                                   <char> <int>
## 1: PRMT_GM1_GMS_MONT_20240807_bc22_D6331     3
## 2: PRMT_GM2_GMS_MONT_20240807_bc23_D6331     3
## 3: PRMT_GM3_GMS_MONT_20240807_bc24_D6331     3
## 4:  PRMT_GO1_GMS_ONT_20240807_bc07_D6331     3
## 5:  PRMT_GO2_GMS_ONT_20240807_bc08_D6331     3
## 6:  PRMT_GO3_GMS_ONT_20240807_bc09_D6331     3
## 7:   PRMT_GP1_GMS_PB_20240807_bc37_D6331     3
## 8:   PRMT_GP2_GMS_PB_20240807_bc38_D6331     3
## 9:   PRMT_GP3_GMS_PB_20240807_bc39_D6331     3

R^2 Barplot

R^2 Lineplot

B-diversity Analysis using sample-wise Bray-Curtis distances

Non-metric Multi-Dimensional Scaling (NMDS)

NMDS v1: color = DNA_isolation_method; shape = Vregion

NMDS v2: color = Vregion; shape = DNA_isolation_method